Cohesin is important for 3D genome organization. Nevertheless, even the complete removal of cohesin has surprisingly little impact on steady-state gene transcription and enhancer activity. Here we show that cohesin is required for the core transcriptional response of primary macrophages to microbial signals, and for inducible enhancer activity that underpins inflammatory gene expression. Consistent with a role for inflammatory signals in promoting myeloid differentiation of hematopoietic stem and progenitor cells (HPSCs), cohesin mutations in HSPCs led to reduced inflammatory gene expression and increased resistance to differentiation-inducing inflammatory stimuli. These findings uncover an unexpected dependence of inducible gene expression on cohesin, link cohesin with myeloid differentiation, and may help explain the prevalence of cohesin mutations in human acute myeloid leukemia.

Three-dimensional organization of the genome has emerged as an important player in transcriptional regulation [1][2][3][4][5][6][7] . In mammals, CTCF and the cohesin complex create sub-megabase structures with elevated internal chromatin contact frequencies, called topologically associating domains (TADs) [8][9][10][11] . Although TADs are generally considered important for transcriptional regulation, ablation of TAD organization by disrupting CTCF or the cohesin complex only caused modest gene expression changes [12][13][14][15] . In contrast, CTCF is required for cell cycle regulation 16 , early embryo development and for the formation of various adult cell types 17 . To uncouple the role of CTCF in cell state transitions and cell proliferation we studied the effect of CTCF be efficiently converted by exogenous CEBPA expression into functional induced macrophages (iMacs) with only one cell division on average (Fig. 1a; Supplementary Note 1) 26 . Using this system, we analyzed a time-series of transdifferentiating cells for genome-wide changes in 3D genome organization (in-situ Hi-C), enhancer activity (ChIP-seq of histone modifications), chromatin accessibility (ATAC-seq) and gene expression (RNA-seq).We first determined genome segmentation into A and B compartments on the basis of the first eigenvector values of a principal component analysis (PCA) on the Hi-C correlation matrix ('PC1 values'). Overall, although most of the genome remained stable, around 14% of A or B compartment regions were dynamic during transdifferentiation, showing transcriptional changed correlating with the altered compartmentalization state (Fig. 1b-f, Extended Data Fig. 1a-d; Supplementary Note 2). Next, we used chromosome-wide insulation potential 27 to identify between 3,100-3,300 TAD borders per time point (Fig. 1g). Boundaries were highly reproducible between biological replicates (Jaccard index>0.99) and enriched in binding sites for CTCF (Extended Data Fig. 1e). Genome-wide insulation scores analysed by PCA over time revealed progressive changes, reflecting a transdifferentiation trajectory (Extended Data Fig. 1f). While 70% of TAD borders were stable across all stages, 18% were lost or gained and 12% were transiently altered (Fig. 1g). CTCF binding was significantly more enriched at stable than at dynamic boundaries (Fig. 1h), as observed earlier 28 . Furthermore, while lost borders showed some CTCF occupancy in B cells that decreased in iMacs, gained borders were depleted for CTCF in both cell states (Fig. 1h), indicating CTCF-independent mechanisms driving local insulation. The dynamic rearrangement of TAD borders during transdifferentiation is illustrated by the DDX54 locus (Fig. 1i), in which a new boundary appears in iMacs without apparent changes in CTCF binding. Furthermore, border gain or loss did not correlate with changes in local gene expression (Extended Data Fig. 1g), indicating that transcription is not a driver of the observed changes. However, whereas motif analysis at ATAC-seq peaks within stable borders indee...

Rhesus macaque (Macaca mulatta) and long-tailed macaque (Macaca fascicularis) are the 2 most commonly used primate model species in biomedical sciences. Although morphological studies have revealed a weak hybridization at the interspecific contact zone, in the north of Indochina, a molecular study has suggested an ancient introgression from rhesus to long-tailed macaque into the Indo-Chinese peninsula. However, the gene flow between these 2 taxa has never been quantified using genetic data and theoretical models. In this study, we have examined genetic variation within and between the parapatric Chinese rhesus macaque and Indo-Chinese long-tailed macaque populations, using 13 autosomal, 5 sex-linked microsatellite loci and mitochondrial DNA sequence data. From these data, we assessed genetic structure and estimated gene flow using a Bayesian clustering approach and the "Isolation with Migration" model. Our results reveal a weak interspecific genetic differentiation at both autosomal and sex-linked loci, suggesting large population sizes and/or gene flow between populations. According to the Bayesian clustering, Chinese rhesus macaque is a highly homogeneous gene pool that contributes strongly to the current Indo-Chinese long-tailed macaque genetic makeup, whether or not current admixture is assumed. Coalescent simulations, which integrated the characteristics of the loci, pointed out 1) a higher effective population size in rhesus macaque, 2) no mitochondrial gene flow, and 3) unilateral and male-mediated nuclear gene flow of approximately 10 migrants per generation from rhesus to long-tailed macaque. These patterns of genetic structure and gene flow suggest extensive ancient introgression from Chinese rhesus macaque into the Indo-Chinese long-tailed macaque population.

Insulators are DNA-protein complexes that play a central role in chromatin organization and regulation of gene expression. In Drosophila different proteins, dCTCF, Su(Hw), and BEAF bind to specific subsets of insulators most of them having in common CP190. It has been shown that there are a number of CP190-binding sites that are not shared with any other known insulator protein, suggesting that other proteins could cooperate with CP190 to regulate insulator activity. Here we report on the identification of two previously uncharacterized proteins as CP190-interacting proteins, that we have named Ibf1 and Ibf2. These proteins localize at insulator bodies and associate with chromatin at CP190-binding sites throughout the genome. We also show that Ibf1 and Ibf2 are DNA-binding proteins that form hetero-oligomers that mediate CP190 binding to chromatin. Moreover, Ibf1 and Ibf2 are necessary for insulator activity in enhancer-blocking assays and Ibf2 null mutation cause a homeotic phenotype. Taken together our data reveal a novel pathway of CP190 recruitment to chromatin that is required for insulator activity.

Cynomolgus macaques (Macaca fascicularis) were introduced on the island of Mauritius between 400 and 500 years ago and underwent a strong population expansion after a probable initial founding event. However, in practice, little is known of the geographical origin of the individuals that colonized the island, on how many individuals were introduced, and of whether the following demographic expansion erased any signal of this putative bottleneck. In this study, we asked whether the current nuclear genome of the Mauritius population retained a signature that would allow us to answer these questions. Altogether, 21 polymorphic autosomal and sex-linked microsatellites were surveyed from 81 unrelated Mauritius individuals and 173 individuals from putative geographical sources in Southeast Asia: Java, the Philippines islands and the Indochinese peninsula. We found that (i) the Mauritius population was closer to different populations depending on the markers we used, which suggests a possible mixed origin with Java playing most probably a major role; and (ii) the level of diversity was lower than the other populations but there was no clear and consistent bottleneck signal using either summary statistics or full-likelihood methods. However, summary statistics strongly suggest that Mauritius is not at mutation-drift equilibrium and favours an expansion rather than a bottleneck. This suggests that on a short time scale, population decline followed by growth can be difficult to deduce from genetic data based on mutation-drift theory. We then used a simple Bayesian rejection algorithm to estimate the number of founders under different demographic models (exponential, logistic and logistic with lag) and pure genetic drift. This new method uses current population size estimates and expected heterozygosity of Mauritius and source population(s). Our results indicate that a simple exponential growth is unlikely and that, under the logistic models, the population may have expanded from an initial effective number of individuals of 10-15. The data are also consistent with a logistic growth with different lag values, indicating that we cannot exclude past population fluctuation.

The homeotic Abdominal-B (Abd-B) gene expression depends on a modular cis-regulatory region divided into discrete functional domains (iab) that control the expression of the gene in a particular segment of the fly. These domains contain regulatory elements implicated in both initiation and maintenance of homeotic gene expression and elements that separate the different domains. In this paper we have performed an extensive analysis of the iab-6 regulatory region, which regulates Abd-B expression at abdominal segment A6 (PS11), and we have characterized two new polycomb response elements (PREs) within this domain. We report that PREs at Abd-B cis-regulatory domains present a particular chromatin structure which is nuclease accessible all along Drosophila development and both in active and repressed states. We also show that one of these regions contains a dCTCF and CP190 dependent activity in transgenic enhancer-blocking assays, suggesting that it corresponds to the Fab-6 boundary element of the Drosophila bithorax complex.